The goal of the work proposed in this fellowship application is to characterize the role of the type 2 deiodinase (D2) in brown adipocyte differentiation and insulin signaling, as well as its regulation by hedgehog signaling. D2 converts thyroxine (T4) to the active form of thyroid hormone 3,5,3'-tri-iodothryonine (T3) in the perinuclear space, thus increasing the supply of T3 to the nucleus. D2 has been shown to be a major determinant of energy expenditure in brown adipose tissue (BAT), and mice with a targeted deletion of D2 (Dio?1') are more susceptible to hypothermia. Preliminary studies demonstrate that D/O2"'" preadipocytes have an impaired ability to differentiate into brown adipocytes. Specifically, Dio2~'~ preadipocytes are insulin resistant, and do not properly undergo mitotic arrest upon terminal differentiation. To determine the mechanism of these defects, I will characterize the protein levels and phosphorylation states of critical factors in insulin/IGF signaling in both the PI3K/Akt and MARK kinase cascades, as well as their dynamic response to both insulin and insulin-like growth factors. I will supplement this research approach by conducting transcriptional profiling of wild type and Dio2"y' preadipocytes to search for less obvious transcriptional defects that might be the cause of either developmental phenotype. As preliminary experiments demonstrate that inhibition of hedgehog signaling promotes brown adipocyte differentiation, I will additionally characterize the role of the hedgehog signaling pathway during differentiation and its regulation of D2 and D3 activity. I anticipate that these studies will provide the mechanisms by which D2-generated T3 control insulin signaling and proliferation during brown adipocyte differentiation, and provide insight into the role of hedgehog signaling in this process. These experiments will give me invaluable research experience in the fields of endocrinology, adipogenesis, and developmental biology that are essential for my future as an independent investigator. Given the diverse functions of D2, it might be an important therapeutic target for the treatment of metabolic diseases such as obesity and type 2 diabetes. [unreadable] [unreadable] [unreadable]